摘要
目的探讨低氧对人骨髓间充质干细胞(hB MSCs)向神经细胞分化的影响。方法将第2代hB M-SCs进行传代、纯化,取第4代hB MSCs进行流式细胞仪检测鉴定;以含1%FBS的α-MEM为基础诱导培养基,根据处理条件不同分为4组:常氧对照组、低氧对照组(3%氧浓度)、常氧诱导组[添加20 ng/mL表皮生长因子(EGF)及20 ng/mL碱性成纤维细胞生长因子(bF GF)]、低氧诱导组(3%氧浓度下添加20 ng/mL EGF及20 ng/mL bF GF)。连续诱导3 d后,采用RT-PCR及Western blot技术检测神经元特异性烯醇化酶(NSE)、微管相关蛋白2(MAP-2)及胶质纤维酸性蛋白(GFAP)在基因及蛋白水平表达变化。结果第4代hB MSCs流式鉴定结果示:骨髓基质标志CD105、CD90、CD29表达分别为95.8%、98.2%、95.1%,而造血细胞标志CD19表达为0.2%;与常氧对照组比较,低氧对照组NSE、MAP-2、GFAP在基因及蛋白水平表达均无明显改变(P>0.05),而常氧诱导组及低氧诱导组相应表达均升高(P<0.05),且低氧诱导组表达高于常氧诱导组(P<0.05)。结论低氧促进EGF联合bF GF对hB MSCs向神经元细胞及神经胶质细胞分化的诱导作用。
Objective To investigate the effect of hypoxia on the differentiation of human bone marrow mesenchy- mal stem cells （hBMSCs） into neural cells. Methods The hMSCs （Passage 2） were purchased from Cyagen Biosei- ences Inc and identified by flow cytometry at Passage 4. The α - MEM medium containing 1% FBS was used as the basal inducing medium. According to culture conditioning, they were divided into 4 groups, normoxia group, hypoxia group （3% 02 ）, normoxia -induced group （adding 20 ng/mL EGF and 20 ng/mL bFGF）, and hypoxia -induced group （ adding 20 ng/mL EGF and 20 ng/mL bFGF under the condition of 3% 02 ）. RT - PCR and Western blot were used to assess the mRNA and protein levels of the neuron markers, including NSE and MAP - 2, and GFAP for glial marker after continuous inducing of 3 days. Results The identification results of flow cytometry showed that the expression of bone marrow stroma marker CDI05, CD90 and CD29 were 95.8%, 98.2% and 95.1%, respectively; while the expression of CD19, as the marker of hematopoietic cell, was 0.2%. Compared to normoxia group, the mRNA and protein levels of NSE, MAP - 2 and GFAP were similar to hypoxic - group, while those were significantly increased both in normoxia - in- duced group and hypoxia - induced group （ P 〈 0. 05 ） ; between which, they were significantly higher in hypoxia - induced group （ P 〈 0. 05 ）. Conclusion Hypoxia promotes the induction of EGF combined with bFGF on differentiation of hBM- SCs into neural cells and neuroglioeytes.
作者
刘礼金
施剑明
殷嫦嫦
LIU Li -jing SHI Jian -ming YIN Chang- chang(Jingdezhen No. 1 People＇s Hospital, Jingdezhen 333000, Jiangxi, China)
出处
《广东医学》
CAS
北大核心
2017年第20期3065-3069,共5页
Guangdong Medical Journal
基金
江西省教育厅科学技术研究项目(编号:GJJ13740)
